Several organic mercurials were found to modify selective aspects of the regulation of adenylate cyclase activity in membranes isolated from fat cells and rat liver. Treatment of the membranes with the mercurials results in irreversible loss of the stimulating actions of GTP and hormones on these systems even after removal of the mercuricals by washing or destruction with thiol reagents. Paradoxically, the activity of the enzyme (seen most dramatically with the fat cell cyclase system) is increased to levels approximating those seen when the enzyme is activated with guanine nucleotides and hormones. Binding of hormones to receptors and the actions of GTP on hormone binding are not altered by organic mercurials, suggesting that the mercurials alter selectively the guanine nucleotide component responsible for mediating hormone action and regulation of adenylate cyclase activity.